Cover member with orientation indicia

ABSTRACT

A cover member for use in the treatment of a sample on a substrate is disclosed. The cover member has fluid flow features and is adapted for use in an instrument, such as a laboratory instrument. The cover member comprises at least one orientation feature detectable by the instrument for ascertaining an orientation of the cover member. An automated method for detecting orientation of a cover member in a sample treatment assembly is also disclosed, in which a processor compares data corresponding to one or more images collected from the sample treatment assembly, with data representing a reference image to determine if a cover member is in the sample treatment assembly.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a National Stage of International Application No.PCT/AU2013/000986 filed Aug. 30, 2013, claiming priority based on U.S.Provisional Application No. 61/696,529 filed Sep. 4, 2012, the contentsof all of which are incorporated herein by reference in their entirety.

FIELD OF THE INVENTION

The present invention relates to a cover member for use in the treatmentof samples such as biological tissue. It relates particularly but notexclusively to a cover member for use with a sample treatment instrumentand which has features that enable the instrument, or a human operatingthe instrument, to determine the orientation of the cover member in theinstrument, including whether or not a cover member is present.

BACKGROUND TO THE INVENTION

Immunohistochemical staining and in situ nucleic acid analysis are toolsused in histological diagnosis and the study of tissue morphology.Immunohistochemical staining involves a series of treatment stepsconducted on a tissue sample mounted on a glass slide to highlight byselective staining, certain morphological indicators of disease states.

Instrumentation and automated sample processing systems exist forautomating steps in sample staining and other treatment processes. Somesystems employ a cover member with physical features which, when placedover the sample on the slide, form a reaction chamber. The cover membermay also have features for controlling the application and transfer offluid reagent within the reaction chamber. Given the high cost ofreagents, the reaction chamber is often very small (e.g. a fewmicroliters in volume) and defined by fine rails on one side of themember. In some cases, these rails are nearly undetectable upon visualinspection, particularly by the inexperienced.

In contrast, deeper contours on the opposing side of the cover member(e.g. to minimise material consumption during manufacture withoutsacrificing strength or durability) are typically easier to see. Thesedeeper contours may mistakenly be identified as defining the reactionchamber by less experienced or unskilled workers, which can lead to thecover member being applied to the slide wrong way up. This can lead toproblems including loss of instrument time, loss of reagent anddeterioration of the sample, to name a few.

It would be desirable to minimise the risk of the cover member beingapplied to the slide in an incorrect orientation (or not at all).

The discussion of the background to the invention included herein,including reference to documents, acts, materials, devices, articles andthe like, is intended to explain the context of the present invention.This is not to be taken as an admission or a suggestion that any of thematerial referred to was published, known or part of the common generalknowledge in the United States of America as at the priority date of anyof the claims.

SUMMARY OF THE INVENTION

It is known to use features such as barcodes or labels on slidescontaining biological samples e.g. to ensure that slides processed by aninstrument are correctly handled when they exit the instrument afterprocessing. However, little or no work has been done by way of employingidentifiable features on a cover member to ascertain the orientation ofthe cover member in the instrument.

Viewed from one aspect, the present invention provides a cover memberfor use in treatment of a sample on a substrate, such as a slide, thecover member having fluid flow features and being adapted for use in aninstrument, wherein the cover member comprises at least one orientationfeature detectable by the instrument for ascertaining an orientation ofthe cover member.

Ascertaining an orientation of the cover member may involve determiningwhether the cover member is correctly oriented in the instrument, orwhether it is incorrectly oriented e.g. inverted and/or rotated.Ascertaining the orientation of the cover member may also involvedetermining automatically the presence or absence of a cover member overa slide in the instrument, prior to commencement of treatment.

An orientation feature may be a machine-readable feature such as a RFIDtag, magnet or the like, or a feature that may be recognised using imagecapturing means and image matching algorithms employed by a processor inthe instrument. Alternatively/additionally, an orientation feature mayinclude a mechanical feature such as a notch or protrusion, pin or thelike which substantially precludes the cover member from beingincorrectly oriented within the instrument or a tray into which multipleslides may be loaded for use in the instrument.Alternatively/additionally, the orientation feature may be identifiableby a human, as is e.g. a word, number, label, image, line, pattern,arrow, physical marking, tactile feature or the like.

In one embodiment, the orientation feature may comprise or beincorporated into the shape of the cover member or a portion thereof.Alternatively/additionally, the orientation feature may be incorporatedinto a fluid input area of the cover member or a nib or other dispensepoint for receiving reagent.

In some embodiments, the orientation feature may comprise a colour orcoloured element wherein, correct orientation of the cover member makesa colour or coloured element on the cover member visible to the operatorand/or detectable by the instrument. By way of example, when the covermember is correctly oriented, the visible surface of the cover member ora portion thereof may be coloured green. Conversely, when incorrectlyoriented a visible surface of the cover member or a portion thereof maybe coloured red to designate incorrect orientation.

The orientation feature may be moulded onto the cover member surface, orit may be embossed, adhered, fused, etched, applied or otherwiseincorporated on or into the cover member surface, or within the covermember material itself.

In one embodiment, the orientation feature is a visible featuredetectable by the instrument, and is substantially circular in shape. Inan embodiment, the orientation feature comprises an annulus on a surfaceof the cover member, the annulus having a substantially semi-circularcross section. The annulus may have any suitable geometry and may belocated anywhere which does not interfere with the other functionalaspects of the cover member, such as a reagent reservoir or nib ontowhich reagent is dispensed. The geometry may be e.g. a radius of about 2mm with feature height of about 0.2 mm although these dimensions are notto be taken as limiting and radii of e.g. 3 mm, 4 mm, 5 mm up to about10 or 11 mm may be acceptable, while feature heights of zero (where thefeature is incorporated into the surface) to about 4 mm may beappropriate, depending on the other features of the cover member andinstrument with which it is to be used.

In a preferred embodiment, an orientation feature on the cover member isdisposed off-centre relative to at least one dimension, preferably acentre-line of the cover member. This ensures that, in the event thatthe cover member is transparent and is wrongly oriented e.g. byinverting around a longitudinal axis, the orientation feature will notbe in the correct location for detection by the instrument.

In an embodiment, the cover member is adapted for use in a sampleholder, such as a slide tray, receivable by the instrument. Here, theorientation feature may be e.g. a protrusion configured to cooperatewith a corresponding recess of the sample holder when the cover memberis correctly oriented in the sample holder. This enables a user toascertain by inspection if the cover member is correctly oriented sincethe cover member will not lie flat on the slide in the sample holderunless the cover member is correctly oriented with the protrusioncooperating with the corresponding recess in the sample holder. Sampleholders may be configured to hold a single sample or a plurality ofsamples such as but not limited to e.g. 3, 5, 10, 15, 20, 30 or moresamples. Since each sample is provided on a slide which must be coveredby an individual cover member, the slide holder has a correspondingnumber of recesses as slides, so that the correct orientation of eachcover member in the sample holder can be ascertained individually.Alternatively/additionally, the sample holder may comprise a protrusionconfigured to cooperate with a corresponding recess of a cover memberwhen the cover member is correctly oriented in the sample holder.Typically, sample holder would have a separate protrusion for each covermember so that the orientation of each cover member in the sample holdercan be ascertained individually.

In one embodiment, the cover member incorporates a magnetic portionthat, when applied in the correct orientation, permits alignment of thecover member in the sample holder. The opposing polarities when thecover member is applied in the incorrect orientation results in adeflection of the cover member or magnetic force of repulsionhighlighting the incorrect orientation the of the cover member.

In one embodiment, the cover member incorporates a magnetic portion suchthat it triggers a sensor, for example, a Hall Effect sensor, whencorrectly oriented. This enables automatic detection of correctorientation (and/or incorrect orientation) by the instrument or a deviceassociated with the instrument of a sample holder,

In one embodiment, the cover member incorporates a portion with a degreeof opacity that triggers a sensor, for example, a photo-diode oroptocoupler, when correctly oriented or conversely, when incorrectlyoriented.

In one embodiment, the orientation feature may incorporate a read/writefunctionality, such as an RFID tag or other such feature that providesthe ability to sense the presence of the cover member, read informationfrom the cover member and/or write information to the feature on thecover member.

In one embodiment, the orientation feature on the cover member mayinclude functionality to provide communication between the cover memberand a controller located in a different location such as onboard theinstrument, connected to the instrument wired/wirelessly or otherwiseconnected. In one embodiment, the controller may be located in a remotegeographical location relative to the cover member.

In one embodiment, cover member may further encode information relevantto the usage of the cover member, including but not limited to, thehistory of the cover member including the number of times the covermember has been used in a staining apparatus, the number of wash cyclesthe cover member has undergone or any other information that may berelevant to the useable life cycle of the cover member.

In an embodiment, the orientation feature may be utilized to calibratethe cover member and/or instrument to compensate for variations in thelaboratory environment. Calibration may provide for correction of theeffect of variables such as lighting, position within the laboratory andother such variable that may impact on the quality of the imaging of theorientation feature.

In an embodiment, an average reference, such as a moving averagereference may be provided to facilitate calibration.Alternatively/additionally, the feature may facilitate calibration ofother aspects of the instrument, such as the dispense point of reagentonto the instrument, for example a nib or other dispense point on thecover member.

Viewed from another aspect, the present invention provides an automatedmethod for detecting orientation of a cover member in a sample treatmentassembly, the method including the steps of: receiving at a processordata representing one or more images collected from the sample treatmentassembly; receiving at the processor data representing a referenceimage; operating the processor to compare the reference image data andthe collected image data to produce a correlation value; and determiningthe presence of a cover member in the sample treatment assembly when thecorrelation value exceeds a threshold value. The reference image datarepresents an image which, if the cover member is correctly oriented,will be detected on the cover member.

In one embodiment, the received data representing collected imagespertains to a plurality of substrates in the sample treatment assemblywhich have been imaged simultaneously. In another embodiment, thereceived data representing collected images pertains to a plurality ofsubstrates in the sample treatment assembly which have been imagedsequentially.

Machine-readable code embodying instructions executable by a processorto perform the methods described herein may also be provided.

BRIEF DESCRIPTION OF THE DRAWINGS

The present invention will now be described in greater detail withreference to the accompanying drawings. It is to be understood that theembodiments shown are examples only and are not to be taken as limitingthe scope of the invention as defined in the claims appended hereto.

FIG. 1 is a schematic illustration of a cover member according to anembodiment of the invention.

FIG. 2 is a schematic illustration showing cover members in a sampleholder, with one of the cover members in an incorrect orientation.

FIG. 3 is an isometric view of part of a sample holder loaded withslides and cover members.

FIG. 3A is an enlarged view of the region A identified in FIG. 3.

FIG. 4 is a flow chart demonstrating steps in an automated method fordetecting presence or absence of a cover member in a treatment assembly,according to an embodiment of the invention.

FIG. 5 is a schematic illustration of an automated instrument with whichthe cover member and/or the method of the present invention may be used.

DETAILED DESCRIPTION

Referring firstly to FIG. 1, there is shown a cover member 100 of thetype that is used for processing a sample a substrate such as a slide300, using an automated instrument 500. An example of an automatedinstrument, with the external casing removed is illustrated in FIG. 5.FIG. 5 shows internal components of the instrument including roboticgantry 502 and probes 504, 506 for dispensing reagent into thedesignated areas of slide staining modules when loaded in theinstrument. The instrument illustrated in FIG. 5 is not yet loaded withslide staining modules in the form of a sample holder prepared with aplurality of slides and respective cover members.

The cover member has fluid control features such as nib 102 onto whichreagent is dispensed for reactions inside reaction chamber 110. In theembodiment illustrated, three orientation features are incorporated: (1)ring 104 which is configured for optical recognition by the instrumentwith which the cover member is used, (2) human and/or machine readablelabel 106 represented by the word “Leica”, and (3) a tab 108 in the formof a projection which is configured to cooperate with a recess in asample holder with which the cover member is used.

FIG. 2 shows a sample holder 200 in the form of a slide tray, whichholds ten slides each identified by reference numeral 300. It is to beunderstood however that the slide holder may hold as few as 1, 2 or 3slides, or as many as 12, 15, 20 etc. The sample holder has slots 201,202, . . . 210 which contain slides 300 and cover members. Slide slots201 to 208 and 210 each contain the cover member 100 in the correctionorientation, ascertainable by the label “Leica” being oriented so thatit can be read from left to right. If the label was inverted, the label“Leica” would not be visible (as it would be facing the slide ratherthan the user) and in the case of transparent cover members, it would bereversed so that on visual inspection, it is obvious that the covermember is not correctly oriented within the sample holder 200. In slot209, it is easily ascertained that the cover member 100 a is incorrectlyorientated because the label “Leica” is not visible.

Furthermore, In FIG. 2 cover member 100 a is not correctly positionedwithin the slot 209 of sample holder 200. This is readily ascertainablebecause cover member 100 a is not in the orientation seen in slots 201to 208 and 210. Cover member 100 a cannot be placed in the sample holder200 because the orientation is incorrect (it has been inverted) and tab108 precludes the cover member 100 a from sitting correctly in the tray.

FIG. 3 provides an isometric view of part of the sample holder in FIG.2, with an encircled region labelled A, providing closer detail of thetab 108. An enlarged version of region A is shown in FIG. 3A which alsoshows recess 218 in the sample holder 200 in which the tab 108 sits whenthe cover member is correctly orientated. In slot 209 of the sampleholder, the tab 108 on the cover member prevents the cover member fromsitting correctly in the slide slot.

In a preferred embodiment, the machine-readable orientation feature 104is a ring configured to be identified by the instrument 500. The featuremay be readily visible to the human operator but need not be. Thus, itmay be replaced by or incorporated into a magnetic, radio frequency,infra-red, or other machine-readable element that can be adapted forautomatic detection by the instrument. In the embodiment illustrated,ring 104 is a substantially circular annular feature that is detectableby the instrument prior to performing a processing run on samples in theinstrument. This enables the instrument system to notify a user that itdetects a cover member is not present on a slide, or the cover memberhas been incorrectly loaded, i.e. in the wrong orientation.

Ideally, the feature 104 is positioned off-centre from the centreline120 of the cover member 100 so that if the cover member is transparentand inverted, the feature 104 will not appear in the image collected bythe instrument. Preferably, the feature is annular with a substantiallysemi-circular cross section. In the embodiment illustrated the annulushas a cross-sectional radius of about 2 mm giving rise to a featureheight of 0.2 mm which is detectable by the instrument and alsoproviding tactile feedback to a user.

The machine-readable feature may be moulded as part of the cover membermaterial, or it may be applied to the surface of the cover member.Preferably, the feature 104 is circular so that the system isunsusceptible to relative rotation between the cover member and theimaging device (not shown) in the instrument.

FIG. 4 is a schematic illustration representing interaction between dataobtained by an imager (such as a scanner or camera in the instrument)and a processor 410 for performing a method for detecting automaticallyan orientation (including presence or absence) of a cover member 100 ina sample treatment assembly such as an instrument 500. The methodincludes receiving at processor 410 data 412 representing one or moreimages collected from the sample treatment assembly as well as data 414representing a reference image. The reference image data represents a“template” of the image that the system is looking for to ascertain theorientation of the cover member. In the embodiment illustrated, this isa ring 104.

The received data representing collected images may contain imagesrepresenting a plurality of slides in the sample treatment assemblywhich have been imaged simultaneously. This may enhance processingefficiency, when automatically ascertaining the orientation of more thanone cover member in the assembly. Alternatively, the received datarepresenting collected images may pertain to a plurality of slides inthe sample treatment assembly, but which have been individually imagedin sequence, rather than simultaneously.

The processor compares reference image data 414 and collected image data412 to produce a correlation value 420. The correlation value representshow well the image represented by the collected data 412 matches thereference image represented by the reference data 410. This may beachieved using software algorithms designed to compare image data e.g.using iterative techniques. A threshold value is stipulated (typicallypre-defined in the algorithm) and when processor 410 determines that thecorrelation value 420 exceeds the threshold 430, a determination can bemade that the cover member 100 is present in the instrument andcorrectly oriented. In the event that the correlation value 420 does notexceed the threshold 430, then a negative result ensues which typicallyprompts an error signal for the user to check the presence andorientation of the cover member.

Outputs from the processor may include e.g. a cropped version of thecollected image and this may be saved to a storage module in theinstrument as evidence of the region that correlated most highly withthe reference image; the x and y coordinates of the detected image mayalso be included. The correlation value may also be stored as well asthe final detection result (i.e. true/false). This data can be used inevent logs generated by the instrument and audit trails aimed atenhancing efficient operation of the instrument and optimising its use.

The automated method may be embodied in machine-readable coderepresenting instructions executable by processor 410 to perform themethod described herein. That code may be updated from time to time e.g.to improve efficiency of image matching algorithms.

Where the terms “comprise”, “comprises”, “comprised” or “comprising” areused in this specification (including the claims) they are to beinterpreted as specifying the presence of the stated features, integers,steps or components, but not precluding the presence of one or moreother features, integers, steps or components or group thereof.

It is to be understood that various modifications, additions and/oralterations may be made to the parts previously described withoutdeparting from the ambit of the present invention as defined in theclaims appended hereto.

It is to be understood that the following claims are provided by way ofexample only, and are not intended to limit the scope of what may beclaimed.

The invention claimed is:
 1. A cover member for use in treatment of asample on a substrate, the cover member having fluid flow features forcontrolling the application and transfer of fluid reagent within areaction chamber defined by the cover member, and being adapted for usein an instrument, wherein the cover member comprises at least oneorientation feature including a human-readable feature also detectableby the instrument for ascertaining absence or presence of the covermember in the instrument, and in the case of presence within theinstrument, for ascertaining an orientation of the cover member, whereinthe at least one orientation feature is incorporated into the covermember, and wherein the at least one orientation feature isasymmetrically disposed off-centre relative to a longitudinal axis ofthe cover member, the longitudinal axis being perpendicular to a shorterand transverse axis of the cover member, and wherein, along thelongitudinal axis, the at least one orientation feature is disposedcloser to a fluid input area of the cover member, at a longitudinal endof the substrate and for receiving the fluid reagent dispensed by theinstrument, than to a longitudinal center of the substrate.
 2. A covermember according to claim 1, wherein the at least one orientationfeature is selected from a group including: a. a machine-readablefeature; b. a mechanical feature; and c. a human-readable feature.
 3. Acover member according to claim 1, wherein the orientation feature issubstantially circular in shape.
 4. A cover member according to claim 1,wherein the ascertainable orientation includes a presence or absence ofthe cover member in the instrument.
 5. A cover member according to claim1, wherein the at least one orientation feature is moulded onto thecover member surface.
 6. A cover member according to claim 3, whereinthe orientation feature comprises an annulus on a surface of the covermember, the annulus having a substantially a semi-circular crosssection.
 7. A cover member according to claim 1, wherein the covermember is adapted for use in a sample holder receivable by theinstrument, and wherein the orientation feature is a protrusionconfigured to cooperate with a corresponding recess of the sample holderwhen the cover member is correctly oriented in the sample holder.
 8. Anautomated method for detecting orientation of a cover member in a sampletreatment assembly, the method including the steps of: a. receiving at aprocessor data representing one or more images collected from the sampletreatment assembly; b. receiving at the processor data representing areference image; c. operating the processor to compare the referenceimage data and the collected image data to produce a correlation value;d. determining the presence of a cover member, having fluid flowfeatures for controlling the application and transfer of fluid reagentwithin a reaction chamber defined by the cover member, in the sampletreatment assembly when the correlation value exceeds a threshold value;and e. providing in the cover member at least one orientation featureincluding a human-readable feature also detectable by the sampletreatment assembly for ascertaining absence or presence of the covermember in the sample treatment assembly, and in the case of presencewithin the sample treatment assembly, for ascertaining an orientation ofthe cover member, the at least one orientation feature beingincorporated into the cover member, wherein the at least one orientationfeature is asymmetrically disposed off-centre relative to a longitudinalaxis of the cover member, the longitudinal axis being perpendicular to ashorter and transverse axis of the cover member, and wherein, along thelongitudinal axis, the at least one orientation feature is disposedcloser to a fluid input area of the cover member, at a longitudinal endof the substrate and for receiving the fluid reagent dispensed by thesample treatment assembly, than to a longitudinal center of thesubstrate.
 9. An automated method according to claim 8 wherein thereceived data representing collected images pertains to a plurality ofsubstrates in the in the sample treatment assembly which have beenimaged simultaneously.
 10. An automated method according to claim 8,wherein the received data representing collected images pertains to aplurality of substrates in the sample treatment assembly which have beenimaged sequentially.
 11. The cover member according to claim 1, whereinthe at least one orientation feature comprises an RFID tag.
 12. Thecover member according to claim 1, wherein the at least one orientationfeature comprises an infra-red symbol.
 13. The cover member according toclaim 1, wherein the cover member comprises a second orientation featureoverlapping the reaction chamber and comprising at least one wordwritten on the cover member at a second longitudinal end of thesubstrate opposite the longitudinal end.